Mycobacterial cell envelopes consist of numerous high molecular weight lipid molecules, most of which induce diverse host immune responses. Among these molecules, cell wall skeleton (CWS) derived from Mycobacterium bovis BCG has been reported recently to be potent in anticancer immunotherapeutics via host antitumor immunity stimulation.1-6) Also, the structure of CWS derived from M. bovis BCG Pasteur and that of cell walls of M. tuberculosis and other mycobacterial species have been studied since the 1970s and speculated to be unique species-and strain-specific complexes consisting of mycolic acid, a very long branched chain-hydroxyl fatty acid, and arabinogalactan, a very large branched chain heteroglycan and extremely stable peptidoglycan part of the cell walls.7-10) However, their detailed structures have not been clarified, because CWS is a huge polymeric compound that does not dissolve in water or any organic solvent, making it difficult to determine the components of CWS and much more difficult to elucidate its structure-biological activity.As a tumor immunotherapy agent, the clinical efficacy of CWS from Mycobacterium bovis BCG has been recognized in many studies. [11][12][13][14][15][16][17][18][19][20][21][22] These clinical results have prompted us to research into the chemistry, bioactivity, and structureactivity relationship of CWS derived from M. bovis BCG Tokyo 172 in more detail, because no such studies on CWS from M. bovis Tokyo 172 have been undertaken to date.Since the most important first step is to secure purified CWS, we developed preparation methods and analytical methods to produce CWS of sufficient quality for use in these studies. In this paper, we describe an optimized preparation method of CWS from M. bovis BCG Tokyo 172 (hereinafter called SMP-105) on the basis of an earlier report, 1) as well as evaluation methods modified with more sensitive and specific technologies. Also, we report analytical results of SMP-105 and show confirmation that the prepared SMP-105 is of sufficient quality for use in research into the chemistry, bioactivity, and structure-activity relationship of CWS. ExperimentalBacterial Strain and Growth Conditions M. bovis BCG Tokyo 172 (ATCC 35737) was grown at 37°C on Sauton medium as surface pellicles until early stationary phase. Cultivated cells were inactivated at 80°C for 30 min and harvested by centrifugation.Preparation of Cell Wall Skeleton (CWS) CWS was prepared according to an optimized and slightly modified method based on an earlier method . 1) Briefly, about 600 g wet cell mass was suspended in 2000 ml water, and then disrupted with Mini DeBEE (BEE International) at 35 kpsi. The disrupted cells were centrifuged at 6760ϫg, 25°C, for 10 min to remove large debris and undisrupted cells. Then supernatant was centrifuged at 18000ϫg, 25°C, for 1 h to obtain pellet, called whole cell walls (WCW). The yield of WCW from 600 g wet cells was about 190 g. WCW was then incubated with benzonase (Merck Ltd.) at 25°C for 17 h to digest nucleic acids. WCW was washed 5 ...

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Iron losses and acoustic noises of the model transformer under DC-biased magnetization were empirically investigated. To clarify the influence of magnetic properties of transformer core materials, two types of grain oriented electrical steels—high permeability grade (HGO) and conventional grade (CGO)—were used as core materials. Iron losses increased with superimposing DC-bias magnetic field (HDC) in both materials, and the iron loss increment in HGO was larger than that in CGO. Acoustic noises increased with increasing HDC in both materials; however, noises emitted from the core of HGO were smaller than those of CGO.